Total RNA was extracted from isolated mouse LV myocytes utilizing TRIZOL reagent (Invitrogen). cDNA was obtained from 1 µg total RNA using MultiScribe reverse transcriptase kit (Applied Biosystems). Real-time RT-PCR was performed with primers designed using the Vector NTI Advance 11 (Invitrogen) software12 (link),55 (link),64 (link). The sequences of primers are indicated in Supplementary Table 1. The StepOnePlus Real-Time PCR system (Applied Biosystems) was employed for quantitative RT-PCR. In each case, cDNA was combined with Power SYBR Green Master Mix (Applied Biosystems) in a 10 µl reaction. Cycling conditions were as follow: 95°C for 10 min followed by 40 cycles of amplification (95°C denaturation for 15 sec, 60°C annealing and extension for 1 min). The melting curve was then obtained. Ct values were normalized with respect to β-2-microglobulin (β2m). To avoid the influence of genomic contamination, forward and reverse primers for each gene were located in different exons. PCR products were run on 3% agarose/1x TBE gel to confirm the specificity of the reaction. Total RNA extracted from mouse brain and skeletal muscle was employed as controls. For fibrotic markers18 (link), RNA was extracted from LV tissue. Ct values were normalized with respect to hypoxanthine guanine phosphoribosyl transferase (Hprt)18 (link).